EP0728840A2 - D-Sorbitol-Dehydrogenase - Google Patents

D-Sorbitol-Dehydrogenase Download PDF

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Publication number
EP0728840A2
EP0728840A2 EP96102405A EP96102405A EP0728840A2 EP 0728840 A2 EP0728840 A2 EP 0728840A2 EP 96102405 A EP96102405 A EP 96102405A EP 96102405 A EP96102405 A EP 96102405A EP 0728840 A2 EP0728840 A2 EP 0728840A2
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Prior art keywords
ifo
gluconobacter
suboxydans
sorbitol
cerinus
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EP96102405A
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English (en)
French (fr)
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EP0728840A3 (de
EP0728840B1 (de
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Tatsuo Hoshino
Setsuko Ojima
Teruhide Sugisawa
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DSM IP Assets BV
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F Hoffmann La Roche AG
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Publication of EP0728840A3 publication Critical patent/EP0728840A3/de
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/0004—Oxidoreductases (1.)
    • C12N9/0006—Oxidoreductases (1.) acting on CH-OH groups as donors (1.1)
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
    • Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y10S435/00—Chemistry: molecular biology and microbiology
    • Y10S435/8215—Microorganisms
    • Y10S435/822—Microorganisms using bacteria or actinomycetales
    • Y10S435/823—Acetobacter

Definitions

  • the present invention concerns a novel -sorbitol dehydrogenase, a process for producing the same and a process for producing ketoses, especially -sorbose utilizing said enzyme.
  • the novel -sorbitol dehydrogenase (hereinafter referred to as SLDH) provided by the present invention catalyzes the oxidation of -sorbitol to -sorbose.
  • SLDH novel -sorbitol dehydrogenase
  • Enzymes which catalyze the oxidation of -sorbitol to -sorbose are known. J. T. Cummins, T.E. King and V.H. Cheldelin ( J. Biol. Chem. , 224 , 323-329, 1957) reported that the cell free extract of Acetobacter suboxydans (syn. Gluconobacter suboxydans ) contains three enzymes participating in pathways of -sorbitol oxidation. Two of these enzymes catalyzing the oxidation of -sorbitol to -sorbose have been purified.
  • NADP nicotine amide adenine dinucleotide phosphate
  • the SLDH provided by the present invention is clearly distinct from the above two enzymes in the structure of subunit of the enzyme, the molecular weight, substrate specificity and optimum pH.
  • the molecular weight of NADP-dependent -sorbose reductase is 60,000, but that of SLDH provided by the present invention consists of homologous subunit having a molecular weight of 79,000 ⁇ 5,000, and does not require NADP to catalyze the reaction.
  • the SLDH of the present invention oxidizes not only -sorbitol and -mannitol, but also -arabitol and erythritol, the optimum pH is 6.0, and the activity is stable even at pH 8.0.
  • a still further object of the present invention is to provide a process for producing -sorbose utilizing said enzyme SLDH.
  • the novel SLDH of the present invention catalyzes the oxidation of -sorbitol to -sorbose in the presence of an electron acceptor according to the following reaction formula:
  • the enzyme does not utilize oxygen as an electron acceptor. This was affirmed by no catalyzing activity of the enzyme to convert -sorbitol to -sorbose using oxygen as a possible electron acceptor. Furthermore, no oxygen consumption was detected in the reaction mixture as detected by a dissolved oxygen probe.
  • any conventional compound which has the ability to act as an electron acceptor can be utilized in conjunction with the enzyme of this invention.
  • an electron acceptor 2,6-dichlorophenolindophenol (hereinafter referred to as DCIP), phenazine methosulfate (hereinafter referred to as PMS) , ferricyanide or cytochrome c can be used.
  • the enzyme assay was performed as follows.
  • the basal reaction mixture for assaying -sorbitol dehydrogenase activity consisted of 50 m potassium phosphate buffer (pH 6.0), 0.25 m DCIP and 0.325 m PMS, which was prepared just before the assay.
  • a cuvette with a 1-cm light path contained 0.4 ml of the basal reaction mixture, 0.1 ml of 0.4 -sorbitol and enzyme solution with a total volume of 0.51 ml.
  • the reference cuvette contained all components except for the substrate.
  • the reaction was started at 25°C with -sorbitol and the enzyme activity was measured as the initial reduction rate of DCIP at 600 nm.
  • One enzyme unit is defined as the amount of the enzyme that catalyzes the reduction of 1 ⁇ mol of DCIP per minute.
  • the extinction coefficient of DCIP at pH 6.0 was 10.8 m -1 .
  • Substrate specificity of the enzyme was determined using the same enzyme assay method as described above in 1) except that the various substrate solution (100 m ) was used instead of -sorbitol.
  • the results of the measurement are shown in Table 1.
  • -sorbitol, -arabitol, erythritol and glycerol were highly oxidized, and -mannitol and -adonitol were also oxidized, at 49.9 and 66.6% of the reaction rate of -sorbitol.
  • the correlation between the reaction rate of the SLDH and pH was determined using the same enzyme assay method as described above in 1) except that various pH's and buffers were used. The results are shown in Table 2.
  • the enzyme showed optimum pH values of 6.0 - 7.0 and showed the highest activity at pH 6.0.
  • the enzyme was kept standing in buffers of various pHs for 16 hours at 4°C, and then the residual activity was measured using the same enzyme assay method as described above in 1). The results of the measurement are shown in Table 3. Over 60% of the activity remained at pHs between 7.0 and 9.0.
  • Thermostability was tested by incubating the enzyme for 5 minutes at various temperatures in 0.01 potassium phosphate buffer (pH 7.0). The residual activity was measured using the same enzyme assay method as described above in 1), after which the treated enzyme was cooled immediately in ice water. The results are shown in Table 4. The enzyme was stable up to 35°C, and lost about 20, 70 and 90% of its activity after it had been incubated at 40, 50 and 60°C, respectively.
  • Table 4 Temperature stability for the SLDH activity Temperature (°C) Relative activity a) (%) 0 100 20 100 25 106 30 106 35 106 40 82.4 50 29.4 60 11.8 a) : Data are expressed as a percentage of the activity at 20°C.
  • the enzyme activities were measured at temperatures from 20 to 60°C in the same assay method as described above in 1). The results are shown in Table 5.
  • the enzyme showed optimum temperature at from 20 to 40°C and showed the highest activity at 30°C. The decrease of 60 and 70% in its activity was observed at 45 and 50°C, respectively, and no more activity was detected at 60°C.
  • Table 5 Optimum temperature for the SLDH activity Temperature (°C) Relative activity a) (%) 20 82.1 28 83.2 30 100 35 91.7 37 77.0 40 74.8 45 39.9 50 28.9 55 4.5 60 0 a) : Data are expressed as a percentage of the activity at 30°C.
  • the apparent Michaelis constant was calculated to be 18 m with DCIP as an electron acceptor for the reaction.
  • the molecular weight of the native SLDH was determined by HPLC using a size exclusion gel column at 280 nm and a flow rate of 1.0 ml per minute.
  • the purified SLDH consisted of homologous subunit with a molecular weight of 79,000 ⁇ 5,000 in the presence of sodium dodecyl sulfate (SDS).
  • Purification of the SLDH is effected by the combination of known purification methods such as ion exchange chromatography, liquid chromatography, adsorption chromatography, gel-filtration chromatography, gel-electrophoresis, salting out and dialysis.
  • the SLDH provided by the present invention can be prepared by cultivating an appropriate microorganism, disrupting the cells and isolating and purifying it from cell free extract of disrupted cells, preferably from the membrane fraction of the microorganism.
  • microorganisms used for the present invention are microorganisms belonging to genus Gluconobacter and Acetobacter . Mutants and variants of said microorganism can be also used in the present invention.
  • strains most preferably used in the present invention are Gluconobacter albidus IFO 3250, Gluconobacter albidus IFO 3251, Gluconobacter albidus IFO 3253, Gluconobacter capsulatus IFO 3462, Gluconobacter cerinus IFO 3263, Gluconobacter cerinus IFO 3264, Gluconobacter cerinus IFO 3265, Gluconobacter cerinus IFO 3267, Gluconobacter cerinus IFO 3270, Gluconobacter dioxyacetonicus IFO 3271, Gluconobacter dioxyacetonicus IFO 3274, Gluconobacter gluconicus IFO 3171, Gluconobacter gluconicus IFO 3285, Gluconobacter gluconicus IFO 3286, Gluconobacter industrius IFO 3260, Gluconobacter melanogenus IFO 3292, Gluconobacter melanogenus IFO 3293
  • microorganisms which can be employed in the process of the present invention, include those which are being preserved in a public depository (culture collection) for delivery to any one upon request such as the Institute of Fermentation Osaka, Japan (IFO).
  • Gluconobacter suboxydans IFO 3255 has been deposited at the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSM), Mascheroder Weg 1b, D-38124 Braunschweig, Germany on February 13, 1995 under the Budapest Treaty. The allotted deposit number is DSM 9715.
  • the microorganism may be cultured in an aqueous medium supplemented with appropriate nutrients under aerobic condition.
  • the cultivation may be conducted at a pH of 3.5 to 8.0, preferably 5.0 to 7.5.
  • the cultivation period varies depending upon the microorganism and the nutrient medium to be used, and is preferably about 6 to 100 hours.
  • a preferred temperature range for carrying out the cultivation is from about 20°C to about 40°C, preferably from about 25°C to about 35°C.
  • the culture medium contains such nutrients as: assimilable carbon sources, such as glycerol, -mannitol, -sorbitol, erythritol, ribitol, xylitol, arabitol, inositol, dulcitol, -ribose, -fructose, -glucose and sucrose, preferably -sorbitol, -mannitol or glycerol; digestible nitrogen sources such as organic substances, for example, peptone, yeast extract, baker's yeast and corn steep liquor, and inorganic substances, for example, ammonium sulfate, ammonium chloride and potassium nitrite; vitamins; and trace elements.
  • assimilable carbon sources such as glycerol, -mannitol, -sorbitol, erythritol, ribitol, xylitol, arabitol, inositol, dulci
  • the SLDH provided by the present invention is useful as a catalyst for the production of -sorbose from -sorbitol.
  • the reaction should be conducted at pH values of about 5.5 to about 8.0, preferably from 6.0 to 7.0, in the presence of an electron acceptor, for example, DCIP, PMS, ferricyanide, cytochrome c and the like in a solvent such as phosphate buffer, tris buffer, citrate buffer and the like.
  • a preferred temperature range for carrying out the reaction is from about 20°C to about 50°C, preferably from 20°C to 40°C. When the pH and temperature are set at about 6.5 to 7.5 and 30°C, respectively, the reaction usually gives particularly good results.
  • the -sorbitol is completely converted to -sorbose within about 2 to 24 hours, preferably within about 12 hours.
  • concentration of -sorbitol in a solvent varies depending upon other reaction conditions, but in general is suitably from about 10 to about 300 g/L, most preferably from about 10 to about 200 g/L.
  • the cultured cells are also useful for the production of ketoses from polyols, especially for the production of -sorbose from -sorbitol.
  • -Sorbose produced from -sorbitol in a solvent is isolated by a combination of such conventional methods as centrifugation and concentration.
  • the solvent containing -sorbose without the isolation step can also be used as the starting material in the industrial production of vitamin C by the Reichstein method.
  • the enzyme may also be used in an immobilized state with an appropriate carrier. Any means of immobilizing an enzyme generally known in the art may be used.
  • the enzyme may be bound directly to a membrane, granules or the like of a resin having functional group(s), or it may be bound to the resin through bridging compounds having bifunctional group(s), for example, glutaraldehyde.
  • Gluconobacter suboxydans IFO 3255 (DSM 9715) was supplied by the Institute for Fermentation, Osaka (IFO), and used throughout this study.
  • the medium consisted of 20 g of -sorbitol, 3 g of yeast extract, 3 g of beef extract, 3 g of corn steep liquor, 10 g of polypeptone, 1 g of urea, 1 g of KH 2 PO 4 , 0.2 g of MgSO 4 ⁇ 7H 2 O, and 1 g of CaCO 3 in 1 liter of deionized water.
  • the pH was adjusted at 7.0 with sodium hydroxide before the addition of CaCO 3 .
  • the cultivation in a flask was carried out aerobically with rotary shaking for one day, or that in a 30-liter jar fermentor was carried out for 21.5 hours at 30°C, 500 rpm for agitation and 15 L/min. for aeration.
  • the broth was centrifuged at 400 x g for 10 minutes to remove calcium carbonate, and then at 10,000 x g to pellet the cells.
  • the cell cake was washed once with physiological saline.
  • the intact cells 200 g wet weight
  • the cells were frozen at -20°C until used.
  • the cells (100g wet weight) were suspended in 200 ml of 50 m phosphate buffer (pH 7.0) and passed through a French pressure cell press at 20,000 psi . After centrifugation to remove intact cells, the supernatant (cell free extract) was centrifuged at 80,000 x g for one hour and this precipitate was designated as the membrane fraction (2.28 g wet weight).
  • the SLDH was isolated from the membrane fraction of Gluconobacter suboxydans IFO 3255 (DSM 9715).
  • DSM 9715 The solubilization was performed by treating the membrane with 0.01 sodium acetate buffer (pH 5.0) containing 1% Triton X-100, 0.1 KCl, 0.1 -sorbitol and about 10 mg/ml of the membrane protein for 2 hours at 5°C.
  • 0.01 sodium acetate buffer (pH 5.0) containing 1% Triton X-100, 0.1 KCl, 0.1 -sorbitol and about 10 mg/ml of the membrane protein for 2 hours at 5°C.
  • the SLDH activity was not recovered from the membrane fraction and the whole activities in both solubilized supernatant and residual membrane fraction were lost under the above conditions.
  • solubilization conditions such as pH value, the concentration of buffer, detergents and KCl
  • the recovery of SLDH activity was 74% into solubilized supernatant from the membrane fraction, when the membrane fraction was mixed in 0.05 potassium phosphate buffer (pH 7.0) containing 1% Triton X-100 and 0.04 -sorbitol for 2 hours at 5°C as shown in Table 8.
  • the enzyme was not solubilized with n -octyl- ⁇ - -glucopyranoside and the activity was lost by the addition of 0.1 KCl.
  • Example 1- To give the active fraction of SLDH for the isolation step ⁇ Example 1- (4) ⁇ from the membrane fraction of Gluconobacter suboxydans IFO 3255 (DSM 9715), frozen membranes were thawed and suspended in the buffer (pH 7.0) to give about 10 mg/ml of protein, and then 1% Triton X-100 and 0.1 -sorbitol were added. The suspension was shaken at 180 rpm for 2 hours, and then centrifuged at 80,000 x g for 60 minutes to remove the precipitate. The SLDH activity was recovered in the solubilized supernatant (200 ml).
  • Example 1-(3) The solubilized supernatant (200 ml) obtained Example 1-(3) was put on a column of DEAE-cellulose (2.5 x 30 cm) equilibrated and washed with the buffer (pH 7.0) containing 0.05 -sorbitol and 0.1% Triton X-100. Elution of the enzyme was performed with 0.1 NaCl in the same buffer. The fractions having enzyme activity were collected.
  • the pooled enzyme fractions (125 ml) from the previous step were dialyzed against two batches of one liter of the buffer containing 0.05M -sorbitol and 0.1% Triton X-100, and put on a DEAE-Sepharose column (1.5 x 50 cm) equilibrated and washed with the same buffer, and the SLDH activity was eluted with a linear gradient of NaCl (0 to 0.2 ).
  • Major enzyme activity was eluted at NaCl concentration ranging from 0.16 to 0.18 .
  • the pooled active fraction (40 ml) from the previous step was dialyzed against two batches of 500 ml of the buffer containing 0.05 -sorbitol and 0.1% Triton X-100.
  • a part of the enzyme (5 ml) was put on a hydroxylapatite column (2.5 x 20 cm) equilibrated. The enzyme activity was eluted during the washing of the column. After the same preparation had been repeated, fractions having enzyme activity were collected. The total volume was 52 ml after the active fraction was dialyzed against the buffer. Then the fraction was concentrated to 10 ml by ultrafiltration (PM10, Amicon).
  • a portion of the enzyme fraction (2 ml) from the previous step was put on a Sephacryl HR300 column (1 x 120 cm) equilibrated with the buffer (pH 7.0) containing 0.05 NaCl, 0.05 -sorbitol and 0.1% Triton X-100 and developed. This fractionation step was repeated and the active fraction was combined. The active fraction dialyzed against the buffer (13 ml) was pooled and stored at -80°C.
  • the purified enzyme with a specific activity of 45.43 units per mg of protein (0.2 mg/ml) was used for the following analysis.
  • the molecular weight of the native -sorbitol dehydrogenase was determined by HPLC (detection, 254 ⁇ m; flow rate, 1 ml/min) using a size exclusion gel column (TSK gel G3000 SW column, 7.8 by 300 mm) equilibrated with 0.1 potassium phosphate buffer (pH 7.0) containing 0.3 NaCl.
  • the molecular weight standards cyanocobalamin (1.35 K), myoglobin (17 K), ovalbumin (44 K), ⁇ -globulin (158 K) and thyroglobulin (670 K) were used.
  • the purified enzyme showed a single peak and the molecular weight was determined to be about 800,000 ⁇ 50,000.
  • the enzyme In the presence of sodium dodecyl sulfate (SDS), the enzyme showed a single band with a molecular weight of about 79,000 ⁇ 5,000. From these results, the purified SLDH consisted of ten homologous subunits.
  • the reaction mixture (1 ml) containing 0.04 each of -sorbitol, -mannitol, -arabitol, erythritol, -adonitol and glycerol, and 8 m PMS was incubated for 4 hours at 30°C in 0.2 potassium phosphate buffer (pH 7.0) with 2.0 units of the purified enzyme.
  • the reaction product was analyzed by HPLC and thin layer chromatography.
  • reaction mixture (total volume 1.04 ml) containing 0.2 ml of purified SLDH ( 0.04 mg protein), 0.04 ml of 0.2 PMS, 0.1 ml of 0.4 -sorbitol, 0.4 ml of 0.5 potassium phosphate buffer (pH 7.0) and 0.3 ml of water was incubated at 30°C with gentle shaking. As a result, -sorbose was formed at the rate of about 1.3 mg/hour.
  • Example 3 Distribution of membrane-bound -sorbitol dehydrogenase
  • nitrocellulose membrane was treated by using the Bio-Rad Immun-Blot kit for Goat Anti-Rabbit, and it was investigated as to which microorganism showed the positive band at the position of molecular weight (MW) 79,000 ⁇ 1,000.
  • MW molecular weight
  • Table 10 all of the tested Gluconobacter strains and Acetobacter aceti subsp. orleansis IFO 3259, Acetobacter aceti subsp. xylinum IFO 3288 and Acetobacter aceti xylinum IFO 13772 showed the positive band.
  • the cell homogenate of Acetobacter aceti subsp. aceti IFO 3281and Acetobacter liquefaciens IFO 12388 showed weakly positive band at the position of MW 79,000 ⁇ 1,000.

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EP96102405A 1995-02-27 1996-02-17 D-Sorbitol-Dehydrogenase Expired - Lifetime EP0728840B1 (de)

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EP96102405A EP0728840B1 (de) 1995-02-27 1996-02-17 D-Sorbitol-Dehydrogenase

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EP95102748 1995-02-27
EP95102748 1995-02-27
EP96102405A EP0728840B1 (de) 1995-02-27 1996-02-17 D-Sorbitol-Dehydrogenase

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EP0728840A2 true EP0728840A2 (de) 1996-08-28
EP0728840A3 EP0728840A3 (de) 2001-12-19
EP0728840B1 EP0728840B1 (de) 2007-04-25

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US (1) US5747301A (de)
EP (1) EP0728840B1 (de)
JP (1) JP3874032B2 (de)
CN (1) CN1136310C (de)
AT (1) ATE360685T1 (de)
DE (1) DE69637038T2 (de)
ES (1) ES2284162T3 (de)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0955358A2 (de) * 1998-03-13 1999-11-10 F. Hoffmann-La Roche Ag Genetisch veränderte, L-Sorbose Reduktase defiziente Mutanten
WO2000049133A1 (de) * 1999-02-19 2000-08-24 Basf Aktiengesellschaft Verfahren zur herstellung von l-sorbose
US6127156A (en) * 1997-08-21 2000-10-03 Roche Vitamins Inc. D-sorbitol dehydrogenase gene
DE19963126A1 (de) * 1999-12-24 2001-07-12 Suedzucker Ag Verfahren zur Herstellung von 6-O-alpha-D-Glucopyranosyl-D-sorbit
US6664082B1 (en) 1998-03-13 2003-12-16 Roche Vitamins Inc. Genetically engineered L-sorbose reductase-deficient mutants
WO2016062232A1 (en) * 2014-10-21 2016-04-28 Dsm Jiangshan Pharmaceutical (Jiangsu) Co., Ltd. A fermentation production process of l-sorbose with high concentration

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CA2365092A1 (en) * 1999-03-17 2000-09-21 Fujisawa Pharmaceutical Co., Ltd. Sorbitol dehydrogenase, gene encoding the same and use thereof
US6204040B1 (en) 1999-04-22 2001-03-20 Korea Research Institute Of Bioscience And Biotechnology Gluconobacter suboxydans sorbitol dehydrogenase, genes and methods of use thereof
JP4775258B2 (ja) 2004-03-17 2011-09-21 味の素株式会社 Ｌ−フクロースの製造方法およびｌ−フコースの製造方法
ES2324128A1 (es) * 2005-09-29 2009-07-30 Proyecto De Biomedicina Cima, S.L. Metodo para el diagnostico de carcinoma hepatocelular mediante el empleo de marcadores moleculares.
KR20140046691A (ko) * 2012-10-10 2014-04-21 주식회사 종근당바이오 6-하이드록시에틸 아미노-l-소르보스로의 생물전환 수율이 우수한 신규 글루코노박터 속 ｃｋｄｂｍ ０９０１ 균주
KR101479133B1 (ko) * 2013-04-17 2015-01-05 건국대학교 산학협력단 신규 ｄ－솔비톨 탈수소화효소 및 이를 이용한 ｌ－소르보스의 생산방법
FR3031983A1 (fr) * 2015-01-23 2016-07-29 Ifp Energies Now Procede de transformation selective de polyols biosources en cetoses
CN105154457B (zh) * 2015-09-21 2018-10-12 南京工业大学 一种来源于丁香假单胞菌的山梨醇脱氢酶基因及其应用
CN106434581A (zh) * 2016-09-18 2017-02-22 南京工业大学 一种固定化山梨醇脱氢酶及其固定化方法与应用

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Cited By (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6127156A (en) * 1997-08-21 2000-10-03 Roche Vitamins Inc. D-sorbitol dehydrogenase gene
US6444449B1 (en) 1997-08-21 2002-09-03 Roche Vitamins, Inc. D-sorbitol dehydrogenase gene
US6653115B1 (en) 1997-08-21 2003-11-25 Roche Vitamins, Inc. D-sorbitol dehydrogenase gene
EP0955358A2 (de) * 1998-03-13 1999-11-10 F. Hoffmann-La Roche Ag Genetisch veränderte, L-Sorbose Reduktase defiziente Mutanten
EP0955358A3 (de) * 1998-03-13 2000-03-15 F. Hoffmann-La Roche Ag Genetisch veränderte, L-Sorbose Reduktase defiziente Mutanten
US6664082B1 (en) 1998-03-13 2003-12-16 Roche Vitamins Inc. Genetically engineered L-sorbose reductase-deficient mutants
KR100710948B1 (ko) * 1998-03-13 2007-04-24 디에스엠 아이피 어셋츠 비.브이. 유전공학적으로 제조된 엘-소르보스 환원효소-결핍 변이주
WO2000049133A1 (de) * 1999-02-19 2000-08-24 Basf Aktiengesellschaft Verfahren zur herstellung von l-sorbose
DE19963126A1 (de) * 1999-12-24 2001-07-12 Suedzucker Ag Verfahren zur Herstellung von 6-O-alpha-D-Glucopyranosyl-D-sorbit
US7208301B2 (en) 1999-12-24 2007-04-24 Südzucker Aktiengesellschaft Mannheim/Ochsenfurt Method for producing 6-0-α-D-glucopyranosyl-D-sorbitol
WO2016062232A1 (en) * 2014-10-21 2016-04-28 Dsm Jiangshan Pharmaceutical (Jiangsu) Co., Ltd. A fermentation production process of l-sorbose with high concentration

Also Published As

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EP0728840A3 (de) 2001-12-19
JPH08242850A (ja) 1996-09-24
DE69637038T2 (de) 2007-12-27
ES2284162T3 (es) 2007-11-01
US5747301A (en) 1998-05-05
DE69637038D1 (de) 2007-06-06
ATE360685T1 (de) 2007-05-15
CN1136310C (zh) 2004-01-28
CN1141341A (zh) 1997-01-29
EP0728840B1 (de) 2007-04-25
JP3874032B2 (ja) 2007-01-31

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